Increased albumin excretion is an indicator of renal damage in a wide variety of conditions, including hypertension, diabetes and aging. Correctly understanding the pathogenesis of albuminuria is prerequisite for interpreting its significance in different conditions, and for designing appropriate therapies. One current view of how the separates small and large molecules assumes that the major physical barrier to macromolecules is the podocyte slit diaphragm (SD) with the glomerular basement membrane (GBM) acting as a coarse pre-filter. Another view ascribes glomerular selectivity to the presence in the GBM of pores of different sizes. In 2003, I proposed a gel permeation/diffusion hypothesis and computer simulation postulating that the GBM is the predominant source of the size selectivity of the kidney, because there is limited space in the GBM (a concentrated gel) into which macro-molecules can permeate. The glomerular filtration rate (GFR) clearly governs all trans-glomerular water transport, but calculations by me and others indicate that diffusion rather than filtration governs Specific Aim (i) will test the primary assumption of the permeation/diffusion hypothesis by directly determining with transmission electron microscopy whether nanoparticles and nanoparticle-tagged proteins of different sizes reach the concentrations in the GBM predicted by a well proven gel permeation equation. Initial experiments will be in vitro with purified GBM;later experiments will be in animals. To test the diffusion postulate we will evaluate the renal distribution of tagged proteins when the renal artery is temporarily clamped following a protocol comparable to that used by Ryan &Karnovsky (1976). Specific Aim (ii) will test the prediction that albuminuria will develop when GFR is substantially decreased. Our preliminary data show that in normal mice angiotensin converting enzyme and/or cyclooxygenase inhibitors in high dose cause temporary reversible albuminuria, exacerbated by a low salt diet. We will carry out experiments to determine the factors leading to this albuminuria, and whether it can be achieved by a decrease in GFR without damaging the GBM, podocytes or tubules. PUBLIC HEALTH RELEVANCE The occurrence of albumin in the urine of an individual (albuminuria) indicates that something is wrong with the kidney. Understanding what changes in the kidney lead to albuminuria is therefore essential for designing proper treatments. We have recently advanced a novel hypothesis that challenges current views on the causes of albuminuria, and are proposing experiments to test this hypothesis.